Ischemic stroke is a major health problem in the U.S. Estrogen is neuroprotective in experimental stroke. Clinical trials of hormone replacement (HR) have shown worse outcomes in patients randomized to HR. Although the adverse effects of HR may be due to progesterone, few effective therapies for stroke are available;therefore, it is critical to delineate the downstream effectors of estrogen mediated (EM) neuroprotection in order to maximize protective effects while minimizing harmful side effects. We have linked estrogen treatment to modulation of angiopoietin-1 (ang-1) mRNA and augmentation of cerebral capillary density in rodent brain. This application proposes to study the consequences of EM ang-1 mRNA augmentation in brain and the effects of therapeutic cerebral angiogenesis with ang-1 and vascular endothelial growth factor (VEGF) in an experimental stroke model. Specific Aims: 1) To delineate the consequences of EM ang-1 mRNA expression in brain by: a) evaluating the localization of ang-1 mRNA and protein expression;b) quantifying ang-1 protein;c) determining the effect of ang-1 on cell signaling;d) evaluating the vascular network's capillary anatomy and permeability, endothelial cell proliferation, apoptosis, and anastomotic architecture. 2) To determine if blocking ang-1 function in vivo attenuates EM neuroprotection. 3) To determine if chronic pre-ischemic treatment with ang-1, or ang-1 with VEGF, alters the microvascular network and offers stroke protection. Methods: Ovariectomized female rats treated with placebo or estrogen and male rats will be used. Gene delivery will be accomplished using adenoviral vectors and intracerebral, as well as intravenous, injection. A transient focal cerebral ischemia model in rats and histological and molecular analyses will be used. Summary: This grant will elucidate the role of estrogen in cerebral angiogenesis and neuroprotection and will investigate therapeutic cerebral angiogenesis as a means of stroke prevention.